Non-Absorbent Articles Containing Additives for Inhibiting the Production of TSST-1

ABSTRACT

Non-absorbent articles, such as tampon applicators, containing an additive are disclosed. The non-absorbent articles include an effective amount of an inhibitory compound, such as thiolactomycin or thiomalonate, for example, to substantially inhibit the production of TSST-1 or exoprotein by Gram positive bacteria.

REFERENCE TO RELATED APPLICATIONS

This application is a divisional application of U.S. patent applicationSer. No. 10/271,457, filed Oct. 16, 2002, which is a patent applicationclaiming the benefit of U.S. Provisional Application Ser. No. 60/331,971and Ser. No. 60/331,937, both of which were filed Nov. 21, 2001. Theentire contents of these applications are incorporated herein byreference.

BACKGROUND OF THE INVENTION

The present invention generally relates to inhibiting the production oftoxic shock syndrome toxin one (TSST-1) by Staphylococcus aureus. Moreparticularly, the present invention relates to inhibiting the productionof TSST-1 in the presence of non-absorbent articles by incorporatingcertain compounds into the absorbent articles having an inhibitoryeffect on Gram positive bacteria and the production of TSST-1.

There exists in the female body a complex process which maintains thevagina and physiologically related areas in a healthy state. In a femalebetween the age of menarche and menopause, the normal vagina provides anecosystem for a variety of microorganisms. Bacteria are the predominanttype of microorganism present in the vagina; most women harbor about 10⁹bacteria per gram of vaginal fluid. The bacterial flora of the vagina iscomprised of both aerobic and anaerobic bacteria. The more commonlyisolated bacteria are Lactobacillus species, Corynebacteria, Gardnerellavaginalis, Staphylococcus species, Peptococcus species, aerobic andanaerobic Streptococcus species, and Bacteroides species. Othermicroorganisms that have been isolated from the vagina on occasioninclude yeast (Candida albicans), protozoa (Trichomonas vaginalis),mycoplasma (Mycoplasma hominis), chlamydia (Chlamydia trachomatis), andviruses (Herpes simplex). These latter organisms are generallyassociated with vaginitis or venereal disease, although they may bepresent in low numbers without causing symptoms.

Physiological, social, and idiosyncratic factors affect the quantity andspecies of bacteria present in the vagina. Physiological factors includeage, day of the menstrual cycle, and pregnancy. For example, vaginalflora present in the vagina throughout the menstrual cycle can includelactobacilli, corynebacteria, ureaplasma, and mycoplasma. Social andidiosyncratic factors include method of birth control, sexual practices,systemic disease (e.g., diabetes), and medications.

Bacterial proteins and metabolic products produced in the vagina canaffect other microorganisms and the human host. For example, the vaginabetween menstrual periods is mildly acidic having a pH ranging fromabout 3.8 to about 4.5. This pH range is generally considered the mostfavorable condition for the maintenance of normal flora. At that pH, thevagina normally harbors numerous species of microorganisms in a balancedecology, playing a beneficial role in providing protection andresistance to infection and makes the vagina inhospitable to somespecies of bacteria such as Staphylococcus aureus (S. aureus). The lowpH is a consequence of the growth of lactobacilli and their productionof acidic products. Microorganisms in the vagina can also produceantimicrobial compounds such as hydrogen peroxide and bactericidesdirected at other bacterial species. One example is the lactocins,bacteriocin-like products of lactobacilli directed against other speciesof lactobacilli.

Some microbial products produced in the vagina may negatively affect thehuman host. For example, S. aureus is a bacteria that commonly colonizeshuman skin and mucous membranes. It causes disease in humans throughinvasion or through the production of toxic proteins. One such diseaseis toxic shock syndrome (TSS), caused by toxic shock syndrome toxin-1(TSST-1) and other similar toxins. When absorbed into the blood stream,TSST-1 produces TSS in non-immune humans.

S. aureus is found in the vagina of approximately 16% of healthy womenof menstrual age. Approximately 25% of the S. aureus isolated from thevagina are found to produce TSST-1. TSST-1 has been identified ascausing TSS in humans.

Symptoms of TSS generally include fever, diarrhea, vomiting and a rashfollowed by a rapid drop in blood pressure. Multiple organ failureoccurs in approximately 6% of those who contract the disease. S. aureusdoes not initiate TSS as a result of the invasion of the microorganisminto the vaginal cavity. Instead as S. aureus grows and multiplies, itcan produce TSST-1. Only after entering the bloodstream does TSST-1toxin act systemically and produce the symptoms attributed to TSS.

Menstrual fluid has a pH of about 7.3. During menses, the pH of thevagina moves toward neutral and can become slightly alkaline. Thischange permits microorganisms whose growth is inhibited by an acidicenvironment the opportunity to proliferate. For example, S. aureus ismore frequently isolated from vaginal swabs during menstruation thanfrom swabs collected between menstrual periods.

When S. aureus is present in an area of the human body that harbors anormal microbial population such as the vagina, it may be difficult toeradicate the S. aureus bacteria without harming members of the normalmicrobial flora required for a healthy vagina. Typically, antibioticsthat kill S. aureus are not an option for use in catamenial productsbecause of their effect on the normal vaginal microbial flora and theirpropensity to stimulate toxin production if all of the S. aureus are notkilled. An alternative to eradication is technology designed to preventor substantially reduce the bacteria's ability to produce toxins.

There have been numerous attempts to reduce or eliminate pathogenicmicroorganisms and menstrually occurring TSS by incorporating intovaginal products one or more biostatic, biocidal, and/or detoxifyingcompounds. For example, L-ascorbic acid has been applied to a menstrualtampon to detoxify toxin found in the vagina. Others have incorporatedmonoesters and diesters of polyhydric aliphatic alcohols, such asglycerol monolaurate, as biocidal compounds (see, e.g., U.S. Pat. No.5,679,369). Still others have introduced other non-ionic surfactants,such as alkyl ethers, alkyl amines, and alkyl amides as detoxifyingcompounds (see, e.g., U.S. Pat. Nos. 5,685,872, 5,618,554, and5,612,045).

Despite the aforementioned attempts, there continues to be a need forcompounds that will effectively inhibit the production of TSST-1 fromGram positive bacteria, and maintain activity even in the presence ofthe enzymes lipase and esterase which can have adverse effects onpotency and which may also be present in the vagina. Further, it isdesirable that the detoxifying compounds useful in the inhibition of theproduction of TSST-1 be substantially non-harmful to the natural florafound in the vaginal area. It is also desirable that the detoxifyingcompound be coated or otherwise introduced onto a non-absorbentsubstrate prior to use.

SUMMARY OF THE INVENTION

The present invention relates to non-absorbent substrates or articleswhich inhibit the production of TSST-1 from Gram-positive bacteria. Thesubstrates are particularly useful for inhibiting the production ofTSST-1 from S. aureus bacteria in the vaginal area. Examples of suitablenon-absorbent substrates which can have the inhibitory compoundsdescribed herein incorporated thereon include non-absorbent incontinencedevices, barrier birth control devices, douches, contraceptive sponges,and tampon applicators. One specific example of a non-absorbentincontinence device is a female barrier incontinence device, such as anincontinence pledget formed from a resilient material like rubber.Another suitable non-absorbent substrate is the applicator used with atampon. For example, the tampon applicator may have one or more of theinhibitory compounds described herein coated on an outer surface, suchthat when the applicator is used to introduce a tampon into a women'svagina the inhibiting compound (typically in the form of a cream, wax,gel or other suitable form) is transferred from the applicator onto thewall of the vagina.

It is a general object of the present invention to provide anon-absorbent article which inhibits the production of TSST-1 from Grampositive bacteria. A more specific object of the present invention is toprovide a non-absorbent incontinence device, a barrier birth controldevice, a contraceptive sponge, tampon applicator, or a doucheincorporating one or more of the inhibitory compounds described hereinwhich act to substantially inhibit the production of TSST-1 by S.aureus.

Another object of the present invention is to provide a non-absorbentsubstrate incorporating one or more inhibitory compounds describedherein in combination with one or more other inhibitory ingredients suchas, but not limited to, for example, aromatic compounds, isoprenoidcompounds, laureth-4, PPG-5 lauryl ether, 1-0-dodecyl-rac-glycerol,disodium laureth sulfosuccinate, glycerol monolaurate,alkylpolyglycosides, polyethylene oxide (2) sorbital ether ormyreth-3-myristate which in combination act to substantially inhibit theproduction of TSST-1 by S. aureus.

A further object of the present invention is to provide a non-absorbentsubstrate that has incorporated thereon one or more compounds that willinhibit the production of TSST-1 from Gram positive bacteria withoutsignificantly imbalancing the natural flora present in the vaginaltract.

Therefore, the present invention is based on the discovery that when oneor more inhibitory compounds are incorporated into or onto anon-absorbent article, the production of TSST-1 in Gram positivebacteria is substantially inhibited. In one embodiment, a firstinhibitory compound has the Structure (I):

wherein: R₃₀₀ is, when present, selected from hydrogen and substitutedor unsubstituted alkyl; R₃₀₁ is selected from the group consisting ofhydrogen, a monovalent, saturated or unsaturated, substituted orunsubstituted hydrocarbyl moiety, and when R₃₀₀ is not present, asubstituted or unsubstituted hydrocarbenyl moiety; R₃₀₂ is selected fromhydrogen, substituted or unsubstituted alkyl, and, R₃₀₃ is selected fromhydrogen, hydroxyl, and alkoxy.

One preferred compound of Structure (I) includes thiolactomycin.

As such, the present invention is directed to a tampon applicatorcomprising a non-absorbent substrate, an effective amount of a firstactive ingredient and an effective amount of a second active ingredient.The first active ingredient has a general formula:

wherein: R₃₀₀ is, when present, selected from hydrogen and substitutedor unsubstituted alkyl; R₃₀₁ is selected from the group consisting ofhydrogen, a monovalent, saturated or unsaturated, substituted orunsubstituted hydrocarbyl moiety, and when R₃₀₀ is not present, asubstituted or unsubstituted hydrocarbenyl moiety; R₃₀₂ is selected fromhydrogen, substituted or unsubstituted alkyl; and, R₃₀₃ is selected fromhydrogen, hydroxy, and alkoxy. The second active ingredient has ageneral formula:R¹⁰—O—R¹¹  (IV)wherein R¹⁰ is a straight or branched alkyl or straight or branchedalkenyl having from 8 to about 18 carbon atoms and R¹¹ is selected fromthe group consisting of an alcohol, a polyalkoxylated sulfate salt and apolyalkoxylated sulfosuccinate salt. The first active ingredient and thesecond active ingredient are effective in substantially inhibiting theproduction of TSST-1 from Gram positive bacteria.

The present invention is further directed to a non-absorbent articlecomprising a non-absorbent substrate, an effective amount of a firstactive ingredient, and an effective amount of a second activeingredient. The first active ingredient has a general formula:

wherein: R₃₀₀ is, when present, selected from hydrogen and substitutedor unsubstituted alkyl; R₃₀₁ is selected from the group consisting ofhydrogen, a monovalent, saturated or unsaturated, substituted orunsubstituted hydrocarbyl moiety, and when R₃₀₀ is not present, asubstituted or unsubstituted hydrocarbenyl moiety; R₃₀₂ is selected fromhydrogen, substituted or unsubstituted alkyl; and, R₃₀₃ is selected fromhydrogen, hydroxy, and alkoxy. The second active ingredient has ageneral formula:R¹⁰—O—R¹¹  (IV)wherein R¹⁰ is a straight or branched alkyl or straight or branchedalkenyl having from 8 to about 18 carbon atoms and R¹¹ is selected fromthe group consisting of an alcohol, a polyalkoxylated sulfate salt and apolyalkoxylated sulfosuccinate salt. The first active ingredient and thesecond active ingredient are effective in substantially inhibiting theproduction of TSST-1 from Gram positive bacteria. The non-absorbentarticle is selected from the group consisting of non-absorbentincontinence devices, barrier birth control devices, and douches.

Other objects and advantages of the present invention, and modificationsthereof, will become apparent to persons skilled in the art withoutdeparture from the inventive concepts defined in the claims.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

In accordance with the present invention, it has been discovered thatcertain compounds as described herein can be incorporated into or onto anon-absorbent article, such as an incontinence pledget, for example, tosubstantially inhibit the production of TSST-1 from Gram positivebacteria. The compounds described herein can also be used in combinationwith surface-active agents such as, for example, compounds with anether, ester, amide, glycosidic, or amine bond linking a C₈-C₁₈ fattyacid to an aliphatic alcohol, polyalkoxylated sulfate salt, orpolyalkoxylated sulfosuccinic salt, to substantially inhibit theproduction of TSST-1 from Gram positive bacteria. Through vigorousresearch and experimentation, it has been discovered that, surprisingly,compounds that inhibit certain fatty acid synthesis routes in bacteriaalso inhibit the production of TSST-1 by S. aureus. Specifically,compounds that inhibit fatty acid II enzymes in other bacterial speciesappear to inhibit their S. aureus homologues.

This invention will be described herein in detail in connection withvarious non-absorbent substrates or products such as non-absorbentincontinence devices, barrier birth control devices, contraceptivesponges, tampon applicators, and douches, but will be understood bypersons skilled in the art to be applicable to other non-absorbentarticles, devices and/or products as well wherein the inhibition ofTSST-1 from Gram positive bacteria would be beneficial. As used herein,the term “non-absorbent article” generally refers to substrates ordevices which include an outer layer formed from a substantiallyhydrophobic material which repels fluids such as menses, blood productsand the like. Suitable materials for construction of the non-absorbentarticles of the present invention include, for example, rubber, plastic,and cardboard.

It has been discovered that certain compounds can substantially inhibitthe production of TSST-1 by Gram positive bacteria and, specifically,the production of TSST-1 from S. aureus bacteria. The inhibitorycompounds useful in the practice of the present invention have thegeneral chemical Structure (I):

wherein: R₃₀₀ when present, is selected from hydrogen or substituted orunsubstituted alkyl (e.g., methyl, ethyl, propyl, etc.); R₃₀₁ isselected from the group consisting of hydrogen, a monovalent, saturatedor unsaturated, substituted or unsubstituted hydrocarbyl moiety (e.g.,methyl, ethyl, etc.); and when R₃₀₀ is not present, a substituted orunsubstituted hydrocarbenyl moiety (e.g., methylene, ethylene, etc.);R₃₀₂ is selected from hydrogen, substituted or unsubstituted alkyl(e.g., methyl, ethyl, propyl, etc.); and, R₃₀₃ is selected fromhydrogen, hydroxyl, and alkoxy (e.g., methoxy, ethoxy, etc.).

In this regard it is to be noted that the hydrocarbyl moieties describedherein include both straight chain and branched chain hydrocarbylmoieties and those interrupted with hetero atoms such as nitrogen,sulfur, and oxygen, for example. One skilled in the art will recognizethat one or more of the inhibitory compounds or structures set forthherein can exist in one or more isomers which are also part of thepresent invention. Also, one or more of the inhibitory compounds setforth herein may exist as salts, which are also part of the presentinvention.

In some embodiments, R₃₀₁ is substituted or unsubstituted oxo, havingfor example the structure:

Alternatively, R₃₀₁ is a monovalent, saturated or unsaturated,substituted or unsubstituted hydrocarbyl moiety having about 4 to about12, or about 6 to about 10, carbon atoms in the main or primary chain(i.e., the longest chain in R₃₀₁ which is attached directly to the ringof Structure (I)). Examples of such moieties include C₄H₄, C₄H₈, C₄H₆,C₈H₁₁, C₈H₁₂, C₈H₁₅, and C₁₂H₁₆, as well as hydrocarbon moieties havingthe following structures:

With respect to Structure (I), an exemplary compound is:

wherein R₃₀₀ and R₃₀₂ are as defined above. One preferred compound ofStructure (I) is thiolactomycin. Another preferred compound of Structure(I) is thiomalonate.

The non-absorbent articles include an inhibitory compound describedherein in an amount effective to substantially inhibit the formation ofTSST-1 when the non-absorbent article is exposed to S. aureus bacteria.Several methods are known in the art for testing the effectiveness ofpotential inhibitory agents on the inhibition of the production ofTSST-1 by S. aureus. One such preferred method is set forth in Example1, below. When tested in accordance with the testing methodologydescribed herein the inhibitory compounds preferably reduce theformation of TSST-1 when the non-absorbent article is exposed to S.aureus by at least about 40%, more preferably by at least about 50%,still more preferably by at least about 60%, still more preferably by atleast about 70%, still more preferably by at least about 80%, still morepreferably by at least about 90%, and still more preferably by at leastabout 95%.

Effective amounts of the inhibitory compounds of Structure (I) capableof significantly reducing the production of TSST-1 are from about 0.05micromoles/gram of non-absorbent product to 5 micromoles/gram ofnon-absorbent product and, desirably, from about 0.1 micromoles/gram ofnon-absorbent product to about 1 micromole/gram of non-absorbentproduct.

Although discussed in the singular, one skilled in the art wouldrecognize that two or more of the inhibitory compounds can be combinedin a non-absorbent article. In such embodiments, it may be possible toreduce the amount of the inhibitory compounds incorporated into theabsorbent article and still achieve satisfactory results.

The inhibitory compounds used in the practice of the present inventioncan be prepared and applied to the non-absorbent article in any suitableform, but are preferably prepared in forms including, withoutlimitation, aqueous solutions, lotions, balms, gels, salves, ointments,boluses, suppositories, and the like. The inhibitory compounds may beapplied to the non-absorbent article using conventional methods. Forexample, the inhibitory compounds described herein can be formulatedinto a variety of formulations such as those employed in currentcommercial douche formulations, or in higher viscosity douches.

The inhibitory compounds as described herein may be employed with one ormore conventional pharmaceutically-acceptable and compatible carriermaterials useful for the desired application. The carrier can be capableof co-dissolving or suspending the compound applied to the non-absorbentarticle. Carrier materials suitable for use in the instant inventioninclude those well-known for use in the cosmetic and medical arts as abasis for ointments, lotions, creams, salves, aerosols, suppositories,gels, and the like.

The non-absorbent articles of the present invention may additionallyinclude adjunct components conventionally found in pharmaceuticalcompositions in their art-established fashion and at theirart-established levels. For example, the non-absorbent articles maycontain additional compatible pharmaceutically active materials forcombination therapy, such as supplementary antimicrobials, antioxidants,anti-parasitic agents, antipruritics, astringents, local anaesthetics,or anti-inflammatory agents.

In another embodiment of the present invention, the inhibitory compoundsof Structure (I) are incorporated into or onto the non-absorbent articlein combination with one or more compounds known to retard TSST-1production without significantly eliminating the beneficial bacterialflora. These include, for example, aromatic compounds, isoprenoidcompounds, compounds with an ether, ester, amide, glycosidic, or aminebond linking a C₈-C₁₈ fatty acid to an aliphatic alcohol,polyalkoxylated sulfate salt, or polyalkoxylated sulfosuccinic salt.

In one embodiment, the compounds of Structure (I) are used incombination with aromatic compounds having the following chemicalstructure:

wherein R¹ is selected from the group consisting of H,

—OR⁵, —R⁶C(O)H, —R⁶OH, —R⁶COOH, —OR⁶OH, —OR⁶COOH, —C(O)NH₂, NH₂ andsalts thereof; R⁵ is a monovalent saturated or unsaturated aliphatichydrocarbyl moiety; R⁶ is a divalent saturated or unsaturated aliphatichydrocarbyl moiety; R⁷ is a trivalent saturated or unsaturated aliphatichydrocarbyl moiety; R⁸ is hydrogen or a monovalent substituted orunsubstituted saturated or unsaturated aliphatic hydrocarbyl moietywhich may or may not be interrupted with hetero atoms; R², R³, and R⁴are independently selected from the group consisting of —H, —OH, C(O)OH,and —C(O)R⁹; R⁹ is a monovalent saturated or unsaturated aliphatichydrocarbyl moiety.

With respect to the aromatic compounds of Structure (II), thehydrocarbyl moieties include both straight chain and branched chainhydrocarbyl moieties and may or may not be substituted and/orinterrupted with hetero atoms. Desirably, the aromatic compounds for usein the present invention contain at least one —OH and/or —C(O)OH group.The —OH and/or —C(O)OH group can be bonded to the aromatic structure, orcan be bonded to an atom which may or may not be directly bonded to thearomatic structure. R⁵ is desirably a monovalent saturated aliphatichydrocarbyl moiety having from 1 to about 15 carbon atoms, preferablyfrom 1 to about 14 carbon atoms. R⁶ is desirably a divalent saturated orunsaturated aliphatic hydrocarbyl moiety having from 1 to about 15carbon atoms, preferably from 1 to about 14 carbon atoms. R⁷ isdesirably a trivalent saturated or unsaturated aliphatic hydrocarbylmoiety having from 1 to about 15 carbon atoms, preferably from 1 toabout 10 carbon atoms, and more preferably from 1 to about 4 carbonatoms. Hetero atoms which can interrupt the hydrocarbyl moiety include,for example, oxygen and sulfur.

Preferred aromatic compounds used in combination with the inhibitorycompounds of Structure (I) include 2-phenylethanol, benzyl alcohol,trans-cinnamic acid, methyl ester of 4-hydroxybenzoic acid,2-hydroxybenzoic acid, 2-hydoxybenzamide, acetyl tyrosine,3,4,5-trihydroxybenzoic acid, lauryl 3,4,5-trihydroxybenzoate,phenoxyethanol, 4-hydroxy-3-methoxybenzoic acid, p-aminobenzoic acid,and 4-acetamidophenol.

The non-absorbent articles of the present invention containing a firstinhibitory compound of Structure (I) combined with a second inhibitoryaromatic compound of Structure (II) contain a sufficient amount of bothinhibitory compounds to substantially inhibit the formation of TSST-1when the absorbent article is exposed to S. aureus bacteria. Preferably,the combination of inhibitory compounds reduces the formation of TSST-1when the non-absorbent article is exposed to S. aureus by at least about40%, more preferably by at least about 50%, still more preferably by atleast about 60%, still more preferably by at least about 70%, still morepreferably by at least about 80%, still more preferably by at leastabout 90%, and still more preferably by at least about 95%.

Generally, the amount of the aromatic compound included in thenon-absorbent article is at least about 0.1 micromoles of aromaticcompound per gram of non-absorbent article, and desirably at least about0.5 micromoles of aromatic compound per gram of non-absorbent articleand preferably about 0.5 micromoles of aromatic compound per gram ofnon-absorbent article to about 100 micromoles of aromatic compound pergram of absorbent article. In a preferred embodiment, the non-absorbentarticle contains from about 1.0 micromole of aromatic compound per gramof non-absorbent article to about 50 micromoles of aromatic compound pergram of non-absorbent article. The amount of first inhibitory compoundof Structure (I) is as described above.

In another embodiment, the inhibitory compounds of Structure (I), arecombined with isoprenoid compounds on the non-absorbent article. As usedherein, the term “isoprenoid compound” means a hydrocarbon structurallybased on multiple isoprene units which may or may not be substituted andmay or may not contain hetero atoms and functional groups such ascarbonyl (e.g., ketones and aldehydes), and hydroxyl (e.g., alcohols).Isoprene, also commonly referred to as 2-methyl-1,3-butadiene, has thefollowing chemical structure:

Desirably, the isoprenoid compounds used in the accordance with thepresent invention are terpene compounds. As used herein, “terpenecompound” refers to compounds which are based on isoprene, but which maycontain heteroatoms such as oxygen and/or hydroxyl (e.g., alcohols), orcarbonyl (e.g., aldehydes and ketones).

Various types of terpenes are useful in accordance with the presentinvention. The terpene compounds may be cyclic or acyclic, and may besaturated or unsaturated. Suitable terpenes include hemiterpenes(terpenes containing 5 carbon atoms), monoterpenes (terpenes containing10 carbon atoms), sesquiterpenes (terpenes containing 15 carbon atoms),diterpenes (terpenes containing 20 carbon atoms), triterpenes (terpenescontaining 30 carbon atoms), tetraterpenes (terpenes containing 40carbon atoms), as well as polyterpenes and mixtures and combinationsthereof. Terpenoids, oxygenated derivatives of terpenes, which may ormay not contain hydroxyl and/or carbonyl groups, are also suitableterpenes.

Examples of monoterpenes useful in the present invention includeα-pinen, β-pinen, campher, geraniol, borneol, nerol, thujone, citral a,limonen, cineole, terpineol, terpinene, terpin (cis and trans),α-myrcene, β-myrcene, dipentene, linalool,2-methyl-6-methylene-1,7-octadiene, and menthol. Examples ofsesquiterpenes useful in the present invention include humulene, ionone,nerolidol and farnesol. An example of a suitable diterpene is phytol. Asuitable triterpene for use in the present invention is squalen.Suitable tetraterpenes for use in the present invention includeα-carotene, β-carotene, γ carotene, δ-carotene, lutein, andviolaxanthin.

Preferred isoprenoid compounds for use in the present invention includeterpineol, β-ionone, terpin (cis and trans), linalool, geraniol,menthol, and mixtures and combinations thereof.

The non-absorbent articles of the present invention containing a firstinhibitory compound of Structure (I) combined with a second inhibitoryisoprenoid contain a sufficient amount of both inhibitory compounds tosubstantially inhibit the formation of TSST-1 when the absorbent articleis exposed to S. aureus bacteria. Preferably, the combination ofinhibitory compounds reduces the formation of TSST-1 when thenon-absorbent article is exposed to S. aureus by at least about 40%,more preferably by at least about 50%, still more preferably by at leastabout 60%, still more preferably by at least about 70%, still morepreferably by at least about 80%, still more preferably by at leastabout 90%, and still more preferably by at least about 95%.

Generally, the amount of the isoprenoid compound included in thenon-absorbent article is at least about 0.1 micromoles of isoprenoidcompound per gram of non-absorbent article, and desirably from about 0.5micromoles of isoprenoid compound per gram of non-absorbent article to100 micromoles of isoprenoid compound per gram of non-absorbent. In apreferred embodiment, the non-absorbent article contains from about 1micromole of isoprenoid compound per gram of non-absorbent article toabout 50 micromoles of isoprenoid compound per gram of non-absorbentarticle. The amount of first inhibitory compound of Structure (I) is asdescribed above.

In another embodiment, the inhibitory compounds of Structures (I) arecombined with certain ether compounds on the non-absorbent substrate.The ether compound has the following chemical structure:R¹⁰—O—R¹¹  (IV)wherein R¹⁰ is a straight or branched alkyl or alkenyl group having achain of from about 8 to about 18 carbon atoms and R¹¹ is selected froman alcohol, a polyalkoxylated sulfate salt or a polyalkoxylatedsulfosuccinate salt.

The alkyl, or the R¹⁰ moiety of the ether compounds useful in thepractice of the present invention can be obtained from saturated andunsaturated fatty acid compounds. Suitable compounds include, C₈-C₁₈fatty acids, and preferably, fatty acids include, without limitation,caprylic, capric, lauric, myristic, palmitic and stearic acid whosecarbon chain lengths are 8, 10, 12, 14, 16, and 18, respectively. Highlypreferred materials include capric, lauric, and myristic acids.

Preferred unsaturated fatty acids are those having one or two cis-typedouble bonds and mixtures of these materials. Suitable materials includemyrystoleic, palmitoleic, linolenic and mixtures thereof.

Desirably, the R¹¹ moiety is an aliphatic alcohol which can beethoxylated or propoxylated for use in the ether compositions incombination with the inhibitory compounds of Structure (I). Suitablealiphatic alcohols include glycerol, sucrose, glucose, sorbitol andsorbitan. Preferred ethoxylated and propoxylated alcohols includeglycols such as ethylene glycol, propylene glycol, polyethylene glycoland polypropylene glycol.

The aliphatic alcohols can be ethoxylated or propoxylated byconventional ethoxylating or propoxylating compounds and techniques. Thecompounds are preferably selected from the group consisting of ethyleneoxide, propylene oxide, and mixtures thereof, and similar ringedcompounds which provide a material which is effective.

The R¹¹ moiety can further include polyalkoxylated sulfate andpolyalkoxylated sulfosuccinate salts. The salts can have one or morecations. Preferably, the cations are sodium, potassium or both.

Preferred ether compounds for use in combination with the inhibitorycompounds of Structure (I) include laureth-3, laureth-4, laureth-5,PPG-5 lauryl ether, 1-0-dodecyl-rac-glycerol, sodium laureth sulfate,potassium laureth sulfate, disodium laureth (3) sulfosuccinate,dipotassium laureth (3) sulfosuccinate, and polyethylene oxide (2)sorbitol ether.

The non-absorbent articles of the present invention containing a firstinhibitory compound of Structure (I) combined with a second inhibitoryether compound contain a sufficient amount of both inhibitory compoundsto substantially inhibit the formation of TSST-1 when the absorbentarticle is exposed to S. aureus bacteria. Preferably, the combination ofinhibitory compounds reduces the formation of TSST-1 when thenon-absorbent article is exposed to S. aureus by at least about 40%,more preferably by at least about 50%, still more preferably by at leastabout 60%, still more preferably by at least about 70%, still morepreferably by at least about 80%, still more preferably by at leastabout 90%, and still more preferably by at least about 95%.

Generally, the amount of ether compound included in the non-absorbentarticle is at least about 0.1 micromoles of ether compound per gram ofnon-absorbent article, and desirably at least about 0.005 millimoles ofether compound per gram of non-absorbent article. In a preferredembodiment, the non-absorbent article contains from about 5.0 micromolesof ether compound per gram of non-absorbent article to about 2millimoles of ether compound per gram of non-absorbent article. Theamount of first inhibitory compound of Structure (I) is as describedabove.

In another embodiment, the inhibitory compounds of Structure (I) arecombined with an alkyl polyglycoside compound on the non-absorbentsubstrate. Suitable alkyl polyglycosides for use in combination with theinhibitory compounds of Structures (I) include alkyl polyglycosideshaving the following chemical structure:H—(Z_(n))—O—R¹⁴  (V)wherein Z is a saccharide residue having 5 or 6 carbon atoms, n is awhole number from 1 to 6, and R¹⁴ is a linear or branched alkyl grouphaving from about 8 to about 18 carbon atoms. Commercially availableexamples of suitable alkyl polyglycosides having differing carbon chainlengths include Glucopon 220, 225, 425, 600, and 625, all available fromHenkel Corporation (Ambler, Pa.). These products are all mixtures ofalkyl mono- and oligoglucopyranosides with differing alkyl group chainlengths based on fatty alcohols derived from coconut and/or palm kerneloil. Glucopon 220, 225, and 425 are examples of particularly suitablealkyl polyglycosides for use in combination with the inhibitorycompounds of Structures (I). Another example of a suitable commerciallyavailable alkyl polyglycoside is TL 2141, a Glucopon 220 analogavailable from ICI Surfactants (Wilmington, Del.).

It should be understood that as referred to herein, analkylpolyglycoside may consist of a single type of alkyl polyglycosidemolecule or, as is typically the case, may include a mixture ofdifferent alkyl polyglycoside molecules. The different alkylpolyglycoside molecules may be isomeric and/or may be alkylpolyglycoside molecules with differing alkyl groups and/or saccharideportions. By use of the term alkyl polyglycoside isomers reference ismade to alkyl polyglycosides which, although including the same alkyether residues, may vary with respect to the location of the alkyl etherresidue in the alkyl polyglycoside as well as isomers which differ withrespect to the orientation of the functional groups about one or morechiral centers in the molecules. For example, an alkyl polyglycoside caninclude a mixture of molecules with saccharide portions which are mono,di-, or oligosaccharides derived from more than one 6 carbon saccharideresidue and where the mono-, di- or oligosaccharide has been etherifiedby reaction with a mixture of fatty alcohols of varying carbon chainlength. The present alkyl polyglycosides desirably include alkyl groupswhere the average number of carbon atoms in the alkyl chain is about 8to about 14 or from about 8 to about 12. One example of a suitable alkylpolyglycoside is a mixture of alkyl polyglycoside molecules with alkylchains having from about 8 to about 10 carbon atoms.

The alkyl polyglycosides employed in the non-absorbent articles incombination with the inhibiting compounds described herein can becharacterized in terms of their hydrophilic lipophilic balance (HLB).This can be calculated based on their chemical structure usingtechniques well known to those skilled in the art. The HLB of the alkylpolyglycosides used in the present invention typically falls within therange of about 10 to about 15. Desirably, the present alkylpolyglycosides have an HLB of at least about 12 and, more desirably,about 12 to about 14.

The non-absorbent articles of the present invention containing a firstinhibitory compound of Structure (I) combined with a second inhibitoryalkyl polyglycoside contain a sufficient amount of both inhibitorycompounds to substantially inhibit the formation of TSST-1 when theabsorbent article is exposed to S. aureus bacteria. Preferably, thecombination of inhibitory compounds reduces the formation of TSST-1 whenthe non-absorbent article is exposed to S. aureus by at least about 40%,more preferably by at least about 50%, still more preferably by at leastabout 60%, still more preferably by at least about 70%, still morepreferably by at least about 80%, still more preferably by at leastabout 90%, and still more preferably by at least about 95%.

Generally, the amount of alkyl polyglycoside compound included in thenon-absorbent article is at least about 0.0001 millimoles of alkylpolyglycoside per gram of non-absorbent article, and preferably at leastabout 0.005 millimoles of alkyl polyglycoside per gram of non-absorbentarticle. In a preferred embodiment, the non-absorbent article containsfrom about 0.005 millimoles per gram of non-absorbent article to about 1millimole per gram of non-absorbent article of alkyl polyglycoside. Theamount of first inhibitory compound of Structure (I) is as describedabove.

In another embodiment, the inhibitory compounds of Structure (I) arecombined with an amide containing compound having the following chemicalstructure:

wherein R¹⁷, inclusive of the carbonyl carbon, is an alkyl group having8 to 18 carbon atoms, and R¹⁸ and R¹⁹ are independently selected fromhydrogen or an alkyl group having from 1 to about 12 carbon atoms whichmay or may not be substituted with groups selected from ester groups,ether groups, amine groups, hydroxyl groups, carboxyl groups, carboxylsalts, sulfonate groups, sulfonate salts, and mixtures thereof.

R¹⁷ can be derived from saturated and unsaturated fatty acid compounds.Suitable compounds include, C₈-C₁₈ fatty acids, and preferably, thefatty acids include, without limitation, caprylic, capric, lauric,myristic, palmitic and stearic acid whose carbon chain lengths are 8,10, 12, 14, 16, and 18, respectively. Highly preferred materials includecapric, lauric, and myristic.

Preferred unsaturated fatty acids are those having one or two cis-typedouble bonds and mixtures of these materials. Suitable materials includemyrystoleic, palmitoleic, linolenic and mixtures thereof.

The R¹⁸ and R¹⁹ moieties can be the same or different and each beingselected from hydrogen and an alkyl group having a carbon chain havingfrom 1 to about 12 carbon atoms. The R¹⁸ and R¹⁹ alkyl groups can bestraight or branched and can be saturated or unsaturated. When R¹⁸and/or R¹⁹ are an alkyl moiety having a carbon chain of at least 2carbons, the alkyl group can include one or more substituent groupsselected from ester, ether, amine, hydroxyl, carboxyl, carboxyl salts,sulfonate and sulfonate salts. The salts can have one or more cationsselected from sodium, potassium or both.

Preferred amide compounds for use in combination with the inhibitorycompounds of Structure (I) include sodium lauryl sarcosinate, lauramidemonoethanolamide, lauramide diethanolamide, lauramidopropyldimethylamine, disodium lauramido monoethanolamide sulfosuccinate anddisodium lauroamphodiacetate.

The non-absorbent articles of the present invention containing a firstinhibitory compound of Structure (I) combined with a second inhibitoryamide compound contain a sufficient amount of both inhibitory compoundsto substantially inhibit the formation of TSST-1 when the absorbentarticle is exposed to S. aureus bacteria. Preferably, the combination ofinhibitory compounds reduces the formation of TSST-1 when thenon-absorbent article is exposed to S. aureus by at least about 40%,more preferably by at least about 50%, still more preferably by at leastabout 60%, still more preferably by at least about 70%, still morepreferably by at least about 80%, still more preferably by at leastabout 90%, and still more preferably by at least about 95%.

The amount of amide-containing compound included in the non-absorbentarticle is at least about 0.0001 millimoles of amide containing compoundper gram of non-absorbent article, and preferably at least about 0.005millimoles of amide containing compound per gram of non-absorbentarticle. In a preferred embodiment, the non-absorbent article containsfrom about 0.005 millimoles per gram of non-absorbent article to about 2millimoles per gram of non-absorbent article. The amount of firstinhibitory compound of Structure (I) is as described above.

In another embodiment, the inhibitory compounds of Structures (I) arecombined with an amine compound having the following chemical structure:

wherein R²⁰ is an alkyl group having from about 8 to about 18 carbonatoms and R²¹ and R²² are independently selected from the groupconsisting of hydrogen and alkyl groups having from 1 to about 18 carbonatoms and which can have one or more substitutional moieties selectedfrom the group consisting of hydroxyl, carboxyl, carboxyl salts andimidazoline.

Desirably, R²⁰ is derived from fatty acid compounds which include,without limitation, caprylic, capric, lauric, myristic, palmitic andstearic acid whose carbon chain lengths are 8, 10, 12, 14, 16, and 18,respectively. Highly preferred materials include capric, lauric, andmyristic. Preferred unsaturated fatty acids are those having one or twocis-type double bonds and mixtures of these materials. Suitablematerials include myrystoleic, palmitoleic, linolenic, and mixturesthereof.

The R²¹ and R²² alkyl groups can further include one or moresubstitutional moieties selected from hydroxyl, carboxyl, carboxylsalts, and R¹ and R² can form an unsaturated heterocyclic ring thatcontains a nitrogen that connects via a double bond to the alpha carbonof the R¹ moiety to form a substituted imidazoline. The carboxyl saltscan have one or more cations selected from sodium potassium or both. TheR²⁰, R²¹, and R²² alkyl groups can be straight or branched and can besaturated or unsaturated.

Preferred amine compounds for use with the inhibitory compounds ofStructures (I) include triethanolamide laureth sulfate, lauramine,lauramino propionic acid, sodium lauriminodipropionic acid, laurylhydroxyethyl imidazonline and mixtures thereof.

In another embodiment, the amine compound can be an amine salt havingthe following chemical structure:

wherein R²³ is an anionic moiety associated with the amine and isderived from an alkyl group having from about 8 to about 18 carbonatoms, and R²⁴, R²⁵, and R²⁶ are independently selected from the groupconsisting of hydrogen and alkyl group having from 1 to about 18 carbonatoms and which can have one or more substitutional moieties selectedfrom the group consisting of hydroxyl, carboxyl, carboxyl salts, andimidazoline. R²⁴, R²⁵, and R²⁶ can be saturated or unsaturated.Desirably, R²³ is a polyalkyloxylated alkyl sulfate. A preferredcompound illustrative of an amine salt is TEA laureth sulfate.

The non-absorbent articles of the present invention containing a firstinhibitory compound of Structure (I) combined with a second inhibitoryamine compound contain a sufficient amount of both inhibitory compoundsto substantially inhibit the formation of TSST-1 when the absorbentarticle is exposed to S. aureus bacteria. Preferably, the combination ofinhibitory compounds reduces the formation of TSST-1 when thenon-absorbent article is exposed to S. aureus by at least about 40%,more preferably by at least about 50%, still more preferably by at leastabout 60%, still more preferably by at least about 70%, still morepreferably by at least about 80%, still more preferably by at leastabout 90%, and still more preferably by at least about 95%.

In accordance with the present invention, the non-absorbent articlecontains an effective amount of the combination of the inhibitorycompounds described herein and amine and/or amine salt compounds. Theamount of amine and/or amine salt compound included in the non-absorbentarticle is at least about 0.00001 millimoles of amine and/or amine saltper gram of non-absorbent article, and preferably at least about 0.0005millimoles of amine and/or amine salt per gram of non-absorbent article.In a preferred embodiment, the non-absorbent article contains from about0.005 millimoles per gram of non-absorbent article to about 2 millimolesper gram of non-absorbent article. The amount of first inhibitorycompound of Structure (I) is as described above.

It will be noted by one skilled in the art that various structures of“R” groups which may be attached to one or more of Structure (I) as setforth herein, are set forth in independent form; that is, they are shownstructurally independent without being directly bound to one of theStructure (I). It is to be noted that the “R” group structures shown inindependent form may have various points of attachment to the mainStructure (I) and that it will be recognized by one skilled in the artwhere appropriate points of attachment can be made on the “R” groups toprovide compounds in accordance with the present invention (some of the“R” groups presented herein having, for example, a dangling orincomplete bond, which is understood to generally indicate where thesestructures will attach to the main Structure (I)).

The present invention is illustrated by the following examples which aremerely for the purpose of illustration and are not to be regarded aslimiting the scope of the invention or manner in which it may bepracticed.

EXAMPLE 1

In this Example, the effect of various test compounds on the growth ofS. aureus and the production of TSST-1 was determined. The testcompound, in the desired concentration (expressed inmicrograms/milliliter) was placed in 10 mL of a growth medium in asterile, 50 mL conical polypropylene tube (Sarstedt, Inc. Newton, N.C.).

The growth medium was prepared by dissolving 37 grams of brain heartinfusion broth (BHI) (Difco Laboratories, Cockeysville, Md.) in 880 mLof distilled water and sterilizing the broth according to themanufacturer's instructions. The BHI was supplemented with fetal bovineserum (FBS) (100 mL) (Sigma Chemical Company, St. Louis, Mo.).Hexahydrate of magnesium chloride (0.021 M, 10 mL) (Sigma ChemicalCompany, St. Louis, Mo.) was added to the BHI-FBS mixture. Finally,L-glutamine (0.027 M, 10 mL) (Sigma Chemical Company, St. Louis, Mo.)was added to the mixture.

Compounds to be tested included hexachlorophene, triclosan and4-hydroxydiphenyl methane. Test compounds were received as solids. Thesolids were dissolved in methanol, spectrophotometric grade (SigmaChemical Company, St. Louis, Mo.) at a concentration that permitted theaddition of 200 microliters of the solution to 10 mL of growth mediumfor the highest concentration tested. Each test compound that wasdissolved in methanol was added to the growth medium in the amountnecessary to obtain the desired final concentration.

In preparation for inoculation of the tubes of growth medium containingthe test compounds, an inoculating broth was prepared as follows: S.aureus (MN8) was streaked onto a tryptic soy agar plate (TSA; DifcoLaboratories Cockeysville, Md.) and incubated at 35° C. The testorganism was obtained from Dr. Pat Schlievert, Department ofMicrobiology, University of Minnesota Medical School, Minneapolis, Minn.After 24 hours of incubation three to five individual colonies werepicked with a sterile inoculating loop and used to inoculate 10 mL ofgrowth medium. The tube of inoculated growth medium was incubated at 35°C. in atmospheric air. After 24 hours of incubation, the culture wasremoved from the incubator and mixed well on a S/P brand vortex mixer. Asecond tube containing 10 mL of the growth medium was inoculated with0.5 mL of the above-described 24 hour old culture and incubated at 35°C. in atmospheric air. After 24 hours of incubation the culture wasremoved from the incubator and mixed well on a S/P brand vortex mixer.The optical density of the culture fluid was determined in a microplatereader (Bio-Tek Instruments, Model EL309, Winooski, Vt.). The amount ofinoculum necessary to give 5×10⁶ CFU/mL in 10 mL of growth medium wasdetermined using a standard curve.

This Example included tubes of growth medium with varying concentrationsof test compounds, tubes of growth medium without test compounds(control) and tubes of growth medium with 20-400 microliters of methanol(control). Each tube was inoculated with the amount of inoculumdetermined as described above. The tubes were capped with foam plugs(Identi-plug plastic foam plugs, Jaece Industries purchased from VWRScientific Products, South Plainfield, N.J.). The tubes were incubatedat 35° C. in atmospheric air containing 5% by volume CO₂. After 24 hoursof incubation the tubes were removed from the incubator and the opticaldensity (600 nm) of the culture fluid was determined and the culturefluid was assayed for the number of colony forming units (CFU) of S.aureus using standard plate count procedures. The remaining culturefluid was prepared for the analysis of TSST-1 as follows: the culturefluid was centrifuged at 2500 rpm at about 2-10° C. for 15 minutes. Thesupernatant was filter sterilized through an Autovial 5 syringelessfilter, 0.2 micrometer pore size (Whatman, Inc., Clifton N.J.). Theresulting fluid was frozen at −70° C. in a Fisherbrand 12×75 millimeterpolystyrene culture tube.

The amount of TSST-1 per mL was determined by a non-competitive,sandwich enzyme-linked immunoabsorbent assay (ELISA). Samples of theculture fluid and the TSST-1 reference standard were assayed intriplicate. The method employed was as follows: four reagents, TSST-1(#TT-606), rabbit polyclonal anti-TSST-1 IgG (LTI-101), rabbitpolyclonal anti-TSST-1 IgG conjugated to horseradish peroxidase(LTC-101), and normal rabbit serum (NRS) certified anti-TSST-1 free(NRS-10) were purchased from Toxin Technology (Sarasota, Fla.). A 10microgram/milliliter solution of the polyclonal rabbit anti-TSST-1 IgGwas prepared in phosphate buffered saline (PBS) (pH 7.4). The PBS wasprepared from 0.016 molar NaH₂PO₄, 0.004 molar NaH₂PO₄—H₂O, 0.003 molarKCl and 0.137 molar NaCl, (Sigma Chemical Company, St. Louis, Mo.). Onehundred microliters of the polyclonal rabbit anti-TSST-1 IgG solutionwas pipetted into the inner wells of polystyrene microplates(Nunc-Denmark, Catalogue Number 439454). The plates were covered andincubated at room temperature overnight. Unbound anti-toxin was removedby draining until dry. TSST-1 was diluted to 10 nanograms/milliliter inPBS with phosphate buffered saline (pH 7.4) containing 0.05% (vol/vol)Tween-20 (PBS-Tween) (Sigma Chemical Company, St. Louis, Mo.) and 1% NRS(vol/vol) and incubated at 4° C. overnight. Test samples were combinedwith 1% NRS (vol/vol) and incubated at 4° C. overnight.

The plates were treated with 100 microliters of a 1% (wt/vol) solutionof the sodium salt of casein in PBS (Sigma Chemical Company, St. Louis,Miss.), covered and incubated at 35° C. for one hour. Unbound BSA wasremoved by 3 washes with PBS-Tween. TSST-1 reference standard (10nanograms/milliliter) treated with NRS, test samples treated with NRS,and reagent controls were pipetted in 200 microliter volumes to theirrespective wells on the first and seventh columns of the plate. Onehundred microliters of PBS-Tween was added to the remaining wells. TheTSST-1 reference standard and test samples were then serially diluted 6times in the PBS-Tween by transferring 100 microliters fromwell-to-well. The samples were mixed prior to transfer by repeatedaspiration and expression. This was followed by incubation for 1.5 hoursat 35° C. and five washes with PBS-T and three washes with distilledwater to remove unbound toxin.

The rabbit polyclonal anti-TSST-1 IgG conjugated to horseradishperoxidase wash diluted according to manufacturer's instructions and 50microliters was added to each microtiter well, except well A-1, theconjugate control well. The plates were covered and incubated at 35° C.for one hour.

Following incubation the plates were washed five times in PBS-Tween andthree times with distilled water. Following the washes, the wells weretreated with 100 microliters of horseradish peroxidase substrate bufferconsisting of 5 milligrams of o-phenylenediamine and 5 microliters of30% hydrogen peroxide in 11 mL of citrate buffer (pH 5.5). The citratebuffer was prepared from 0.012 M anhydrous citric acid and 0.026 Mdibasic sodium phosphate. The plates were incubated for 15 minutes at35° C. The reaction was stopped by the addition of 50 microliters of a5% sulfuric acid solution. The intensity of the color reaction in eachwell was evaluated using the BioTek Model EL309 microplate reader (OD490 nanometers). TSST-1 concentrations in the test samples weredetermined from the reference toxin regression equation derived duringeach assay procedure. The efficacy of the compounds in inhibiting theproduction of TSST-1 is shown in Table I below.

In accordance with the present invention, the data in Table 1 shows thatS. aureus (MN8), when compared to the control, produced significantlyless TSST-1 in the presence of the hexachlorophene and triclosancompounds. At the concentration tested, these compounds reduced theamount of toxin produced by 68% to 88%. Although4-hydroxydiphenyl-methane did reduce the toxin production by about 24%,it lacks the chlorine and hydrogen groups that have been shown tostabilize triclosan in the active site of the enzyme/NAD complex. TABLE1 ELISA: Amount Optical TSST-1 Reduction Test Density ng/OD of CompoundCompound 600 nm CFU/mL unit Toxin (%) Methanol 200 μL 0.569 2.9E+08 1038N/A Hexa- 2 μg/mL 0.350 3.7E+08 330 68% chlorophene Triclosan 0.01 μg/mL0.271 1.0E+08 129 88% 4- 2 μg/mL 0.581 1.1E+08 785 24% Hydroxy-diphenyl- methaneN/A = Not Applicable

EXAMPLE 2

In this Example, the growth of, and TSST-1 production by, S. aureusFRI-1169 and 3 mutants able to grow in the presence of triclosan, wasevaluated. S. aureus FRI-1169 was obtained as a lyophilized culture fromthe stock collection of Merlin Bergdoll (Food Research Institute,Madison Wis.). The mutants were selected by plating overnight growth ofS. aureus FRI-1169 in growth medium onto tryptic soy agar platescontaining 5 micrograms/milliliter triclosan. The effect of triclosanwas determined by placing a range of concentrations, expressed inmicrograms/milliliter, in 10 mL of growth medium as set forth inExample 1. The samples were then tested and evaluated utilizing theprocedure set forth in Example 1. The effect of the triclosan on thegrowth of S. aureus FRI-1169 and on the production of TSST-1 is shown inTable 2.

In accordance with the present invention, the data shows that S. aureusFRI-1169, when compared to the control, produced less TSST-1 in thepresence of triclosan. In addition, mutants selected for their abilityto grow in the presence of triclosan showed a reduction in toxinproduction, compared to the parent strain, of 71%-95% in the presence oftriclosan. TABLE 2 ELISA: Amount Optical TSST-1 Reduction Test Densityng/OD of Compound Compound 600 nm CFU/mL unit Toxin % Methanol 200 μL0.577 1.79E+09 958 N/A Triclosan 0.5 μg/mL 0.625 1.50E+09 40 96% Mutant#1 5 μg/mL 0.530 1.78E+09 47 95% Mutant #2 5 μg/mL 0.464 1.41E+09 11488% Mutant #3 5 μg/mL 0.514 1.58E+09 282 71%N/A = Not Applicable

EXAMPLE 3

In this Example, the growth of, and TSST-1 production by, S. aureusFRI-1187 and 3 mutants able to grow in the presence of triclosan wereevaluated. S. aureus FRI-1187 was obtained as a lyophilized culture fromthe stock collection of Merlin Bergdoll (Food Research Institute,Madison Wis.). The mutants were selected by plating overnight growth ofS. aureus FRI-1187 in growth medium onto tryptic soy agar platescontaining 5 microgram/milliliter triclosan. The effect of triclosan wasdetermined by placing a range of concentrations, expressed inmicrogram/milliliter, in 10 mL of a growth medium as in Example 1. Thesamples were then tested and evaluated as in Example 1. The effect ofthe triclosan on the growth of S. aureus FRI-1187 and mutants and on theproduction of TSST-1 is shown in Table 3 below.

In accordance with the present invention, Table 3 shows that S. aureusFRI-1187, when compared to the control, produced less TSST-1 in thepresence of triclosan. In addition, mutants selected for their abilityto grow in the presence of triclosan showed a reduction in toxinproduction, compared to the parent strain, of 85%-94% in the presence oftriclosan. TABLE 3 ELISA: Amount Optical TSST-1 Reduction Test Densityng/OD of Compound Compound 600 nm CFU/mL unit Toxin % Methanol 200 uL0.594 4.40E+09 675 N/A Triclosan 0.5 ug/mL 0.156 1.56E+09 95 86% Mutant#4 10 ug/mL 0.613 Not 102 85% Deter- mined Mutant #5 10 ug/mL 0.618 Not42 94% Deter- mined Mutant #6 10 ug/mL 0.613 1.41E+09 42 94%N/A = Not Applicable

EXAMPLE 4

In this Example, an experiment was conducted to evaluate the growth of,and TSST-1 production by, S. aureus in the presence of cerulenin. Theeffect of the test compounds was determined by placing the desiredconcentration, expressed in micrograms/milliliter, in 10 mL of a growthmedium as set forth in Example 1. The compounds were then tested andevaluated as in Example 1. The effect of the test compounds on thegrowth of S. aureus MN8 and the production of TSST-1 is shown in Table4.

In accordance with the present invention, the data in Table 4 show thatS. aureus MN8, when compared to the control, produce significantly lessTSST-1 in the presence of cerulenin. At the concentrations tested,cerulenin reduced the amount of toxin produced by 89% to 93% on theconcentration tested. TABLE 4 Amount ELISA: Test Optical TSST-1Reduction Compound Density ng/OD of Compound (ug/mL) 600 nm CFU/mL unitToxin % Methanol 120 uL 0.567 6.6E+08 1088 N/A Cerulenin 120 0.5393.3E+08 123 89% Methanol 80 uL 0.526 3.9E+08 1003 N/A Cerulenin 80 0.6269.1E+08 70 93%N/A = Not Applicable

EXAMPLE 5

In this Example, an experiment was conducted to evaluate the growth of,and TSST-1 production by, S. aureus in the presence of cerulenin. Theeffect of the test compound was determined by placing the desiredconcentration, expressed in percent of the active compound, in 100 mL ofgrowth medium (as described in Example 1) in a 500 mL fleaker (CorningLife Sciences, Acton, Mass.). The fleakers were incubated in a 37° C.gyratory water bath and shaken at 180 rpm. Growth was monitoredperiodically by optical density (600 nm) readings. When the opticaldensity reached approximately 1.0, samples were taken and prepared forELISA testing as described in Example 1. The effect of the testcompounds on the growth of S. aureus MN8 and on the production of TSST-1is shown in Table 5 below.

In accordance with the present invention, the data show that S. aureusMN8, when compared to the control, produced significantly less TSST-1 inthe presence of cerulenin. At the concentration tested, these compoundsreduced the amount of toxin produced by 83% to 95%. TABLE 5 AmountOptical ELISA: Reduction Test Density TSST-1 of Compound Compound 600 nmng/OD unit Toxin % Growth Medium 0 1.008 (5 hr) 1653 N/A Cerulenin 40ug/mL 1.128 (6 hr) 71 95% Cerulenin 20 ug/mL 0.956 (5 hr) 278 83%N/A = Not Applicable

In view of the above, it will be seen that the several objects of theinvention are achieved. As various changes could be made in theabove-described non-absorbent articles without departing from the scopeof the invention, it is intended that all matter contained in the abovedescription be interpreted as illustrative and not in a limiting sense.

1. A tampon applicator comprising a non-absorbent substrate, aneffective amount of a first active ingredient and an effective amount ofa second active ingredient, the first active ingredient having a generalformula:

wherein: R₃₀₀ is, when present, selected from hydrogen and substitutedor unsubstituted alkyl; R₃₀₁ is selected from the group consisting ofhydrogen, a monovalent, saturated or unsaturated, substituted orunsubstituted hydrocarbyl moiety, and when R₃₀₀ is not present, asubstituted or unsubstituted hydrocarbenyl moiety; R₃₀₂ is selected fromhydrogen, substituted or unsubstituted alkyl; and, R₃₀₃ is selected fromhydrogen, hydroxy, and alkoxy, the second active ingredient having ageneral formula:R¹⁰—O—R¹¹  (IV) wherein R¹⁰ is a straight or branched alkyl or straightor branched alkenyl having from 8 to about 18 carbon atoms and R¹¹ isselected from the group consisting of an alcohol, a polyalkoxylatedsulfate salt and a polyalkoxylated sulfosuccinate salt, and wherein thefirst active ingredient and the second active ingredient are effectivein substantially inhibiting the production of TSST-1 from Gram positivebacteria.
 2. The tampon applicator as set forth in claim 1 wherein thefirst active ingredient is thiolactomycin.
 3. The tampon applicator asset forth in claim 1 wherein the first active ingredient is present inan amount of from about 0.05 micromoles/gram of non-absorbent substrateto about 5 micromoles/gram of non-absorbent substrate, and the secondactive ingredient is present in an amount of at least about 0.1micromoles per gram of non-absorbent substrate.
 4. The tampon applicatoras set forth in claim 1 wherein R¹⁰ is a straight or branched alkylgroup.
 5. The tampon applicator as set forth in claim 1 wherein R¹⁰ is astraight or branched alkenyl group.
 6. The tampon applicator as setforth in claim 1 wherein R¹⁰ is obtained from the group consisting ofcaprylic acid, capric acid, lauric acid, myristic acid, palmitic acid,and stearic acid.
 7. The tampon applicator as set forth in claim 1wherein R¹¹ is an aliphatic alcohol.
 8. The tampon applicator as setforth in claim 7 wherein R¹¹ is an aliphatic alcohol selected from thegroup consisting of glycerol, glycol, sucrose, glucose, sorbitol, andsorbitan.
 9. The tampon applicator as set forth in claim 1 wherein thesecond active ingredient is selected from the group consisting oflaureth-3, laureth-4, laureth-5, PPG-5 lauryl ether,1-0-dodecyl-rac-glycerol, sodium laureth sulfate, potassium laurethsulfate, disodium laureth (3) sulfosuccinate, dipotassium laureth (3)sulfosuccinate and polyethylene oxide (2) sorbitol ether.
 10. The tamponapplicator as set forth in claim 1 wherein the first active ingredientand the second active ingredient are effective in substantiallyinhibiting the production of TSST-1 from Staphylococcus aureus bacteria.11. A non-absorbent article comprising a non-absorbent substrate, aneffective amount of a first active ingredient, and an effective amountof a second active ingredient, the first active ingredient having ageneral formula:

wherein: R₃₀₀ is, when present, selected from hydrogen and substitutedor unsubstituted alkyl; R₃₀₁ is selected from the group consisting ofhydrogen, a monovalent, saturated or unsaturated, substituted orunsubstituted hydrocarbyl moiety, and when R₃₀₀ is not present, asubstituted or unsubstituted hydrocarbenyl moiety; R₃₀₂ is selected fromhydrogen, substituted or unsubstituted alkyl; and, R₃₀₃ is selected fromhydrogen, hydroxy, and alkoxy, the second active ingredient having ageneral formula:R¹⁰—O—R¹¹  (IV) wherein R¹⁰ is a straight or branched alkyl or straightor branched alkenyl having from 8 to about 18 carbon atoms and R¹¹ isselected from the group consisting of an alcohol, a polyalkoxylatedsulfate salt and a polyalkoxylated sulfosuccinate salt, wherein thefirst active ingredient and the second active ingredient are effectivein substantially inhibiting the production of TSST-1 from Gram positivebacteria, and wherein the non-absorbent article is selected from thegroup consisting of non-absorbent incontinence devices, barrier birthcontrol devices, and douches.
 12. The tampon applicator as set forth inclaim 11 wherein the first active ingredient is thiolactomycin.
 13. Thetampon applicator as set forth in claim 11 wherein the first activeingredient is present in an amount of from about 0.05 micromoles/gram ofnon-absorbent substrate to about 5 micromoles/gram of non-absorbentsubstrate, and the second active ingredient is present in an amount ofat least about 0.1 micromoles per gram of non-absorbent substrate. 14.The tampon applicator as set forth in claim 11 wherein R¹⁰ is a straightor branched alkyl group.
 15. The tampon applicator as set forth in claim11 wherein R¹⁰ is a straight or branched alkenyl group.
 16. The tamponapplicator as set forth in claim 11 wherein R¹⁰ is obtained from thegroup consisting of caprylic acid, capric acid, lauric acid, myristicacid, palmitic acid, and stearic acid.
 17. The tampon applicator as setforth in claim 11 wherein R¹¹ is an aliphatic alcohol.
 18. The tamponapplicator as set forth in claim 17 wherein R¹¹ is an aliphatic alcoholselected from the group consisting of glycerol, glycol, sucrose,glucose, sorbitol, and sorbitan.
 19. The tampon applicator as set forthin claim 11 wherein the second active ingredient is selected from thegroup consisting of laureth-3, laureth-4, laureth-5, PPG-5 lauryl ether,1-0-dodecyl-rac-glycerol, sodium laureth sulfate, potassium laurethsulfate, disodium laureth (3) sulfosuccinate, dipotassium laureth (3)sulfosuccinate and polyethylene oxide (2) sorbitol ether.
 20. The tamponapplicator as set forth in claim 11 wherein the first active ingredientand the second active ingredient are effective in substantiallyinhibiting the production of TSST-1 from Staphylococcus aureus bacteria.